Bidirectional cross talk between ERα and EGFR signalling pathways regulates tamoxifen-resistant growth

We have previously demonstrated that oestrogen receptor alpha (ER alpha) modulates epidermal growth factor receptor (EGFR)/mitogen-activated protein kinase (MAPK) signalling efficiency in a tamoxifen-resistant MCF-7 breast cancer cell line (Tam-R). In the present study we have investigated whether this cross-talk between EGFR/ MAPK and ER alpha signalling pathways is bidirectional by examining the effects of EGFR/MAPK activity on ER functionality In the same cell line. Elevated expression levels of phosphorylated serine 118 (S118) ER alpha were observed in the Tam-R compared to the parental wild type MCF-7 cell line (WT-MCF-7) under basal growth conditions. Phosphorylation of ER alpha at S118 was regulated by the EGFR/MAPK pathway in Tam-R cells being increased in response to amphiregulin (AR) and inhibited by the selective EGFR tyrosine kinase inhibitor, gefitinib and the MEK1/2 inhibitor, PD184352. Recruitment of the co-activators p68 RNA helicase and SRC1 to ER alpha, oestrogen response element (ERE) activity and Tam-R cell growth were similarly EGFR/ MAPK-regulated. Chromatin immunoprecipitation (ChIP) studies revealed that in Tam-R cells the ER alpha assembled on the AR gene promoter and this was associated with elevated basal expression of AR mRNA. Furthermore, AR mRNA expression was under the regulation of the EGFR/MAPK and ERa signalling pathways. Neutralising antibodies to AR inhibited EGFR/ERK1/2 activity, reduced S118 ER alpha phosphorylation and reduced AR mRNA expression in TAM-R cells. These findings suggest that ER alpha function in Tam-R cells is maintained as a consequence of EGFR/MAPK-mediated phosphorylation at serine residue 118 resulting in the generation of a self-propogating autocrine growth-regulatory loop through the ER alpha-mediated production of AR.